IEDB 3D Viewers (IEDB-3D 2.0)

The IEDB has been enhanced to enable better visualization of 3D structures. We are now using NCBI's iCn3D viewer to visualize the structures - please refer to the help resources available on the NCBI page for general introductions on how to utilize the viewer.  

The IEDB has implemented three custom viewer applications that visualize different aspects of IEDB data:

1. Assay Viewer - Displays an experimentally determined 3D structure from the Protein Data Bank (PDB), which directly shows epitope recognition. Depending on the assay type, it displays an epitope being bound by MHC, an epitope bound by MHC and a TCR, or an epitope bound by an antibody, and provides consistent naming of chains and pre-calculated contacts.
2. Epitope Viewer - Displays where an individual epitope is located in its source antigen, using 3D models of the antigen either pulled from the PDB or the AlphaFold Protein Structure Database.
3. ImmunomeBrowser Viewer - works like the Epitope Viewer but maps a set of epitopes to an antigen and highlights which antigen residues are frequently part of epitopes that are commonly recognized (i.e., immunogenic regions).

We welcome your feedback and comments via email to help@iedb.org.

Each of these three viewers is described in more detail in the following: 

1.  Assay Viewer

The assay viewer is available from the ‘Assays’ tab in the query results for assays that directly generate structural information captured in a PDB structure. If the assay viewer is available, a 3D symbol is displayed next to the assay ID (see #1 in Fig. 1A). To specifically search for assays that have such information available, the user can select the ‘3D structure available’ checkbox in the ‘Epitope’ panel of the Filter Options (see #2 in Fig. 1A). 

Clicking the 3D logo launches the assay viewer (Fig. 1B). Using the IEDB curation data, the specific chains are marked to make the MHC/TCR/Ab and epitope chains selectable (see #1 in Fig. 1B). In addition, the residues on the receptor chains in contact with the epitope can be selected to highlight them (see #2 in Fig. 1B). 

Figure 1A: A print screen from the IEDB showing how to access the 3D structures related to immune receptors. Any assay with any atomic model available can be accessed by clicking the 3D button (#1). The user can filter for assays containing only 3D structures available by selecting the checkbox ‘3D Structure Assays’ (#2).

Figure 1B: An image of the viewer pop-up when the user clicks on the 3D button from the ‘Assays’ tab. iCn3D is displaying a TCR/pMHC structure. Box #1 contains the chain controls, which will select various chains of the PDB, and in box #2, the user can click on the checkbox to display pre-calculated interaction between the chains.

2.  Epitopes Tab:

On the IEDB-3D 2.0, we have also included 3D data on the ‘Epitopes’ tab. To visualize the epitope in a 3D antigen, click on the 3D button (Fig. 2A). It will automatically open the atomic model for that antigen in a new browser tab with the selected epitope region highlighted in green (Fig. 2B).

Figure 2A: An image extracted from the IEDB webpage shows how to access the atomic model available to a specific epitope. Any epitope that contains the 3D button has a PDB crystal or an Alphafold-modeled structure with the epitope highlighted.

Figure 2B: An image extracted from the IEDB webpage showing the new tab that is opened upon selecting the 3D icon in the ‘Epitopes’ results tab. iCn3D displays the selected epitope in green highlight on the antigen’s structure. This is an example of when the PDB structure is available.

3.  ImmunomeBrowser (IB) 3D Viewer:

To access our third 3D viewer via the ImmunomeBrowser, users can navigate to the 'Antigens' tab on the IEDB results page and select the histogram-like symbol, which will open the ImmunomeBrowser (Fig. 3A). The red arrow indicates where the IB will be opened from. You can read up on the ImmunomeBrowser in this help article. 

Figure 3A: Screenshot of the IEDB's results page when the user has navigated to the 'Antigens' tab. The ImmunomeBrowser can be accessed by clicking on the histogram-like symbol.

Once the IB has opened, the user will find a panel on the right side of the page where they can access a dropdown menu (Fig. 3B). Here, the IB 3D viewer will be opened within the same panel and will display epitopes mapped to a PDB structure. Users will be able to select their structure of choice drop the dropdown menu. If there are more than five 3D structures available for that same antigen, the results will be filtered, and only five will be displayed in the menu based on:

1. The highest total number of epitopes mapped on that crystal
2. Best resolution

Our filtering process can also include any other PDB structures that are of particular interest to researchers based on feedback from our group or the Help Center. 

Figure 3B: Screenshot of the ImmunomeBrowser, where a Dropdown menu is available on the IB page, showing the PDB or Alphafold ID and the number of epitopes mapped into that atomic model. Users can select their structure of choice to be displayed.

After selecting a 3D structure to be displayed, the rows in the epitope table will be colored yellow if that epitope is mapped on that structure or white if it is not (Fig. 3C). The user can also choose between response frequency ('Map RF' button), and response frequency normalized ('Map RF (Normalized)' button), and both functions will color the atomic model based on its response frequency. Therefore, regions with more immunogenic epitopes will be displayed in dark blue, and regions with no immunogenicity will be displayed in white. Light blue indicates regions with low immunogenicity.

Figure 3C: A print screen from the IB page shows the response frequency and epitope assay count graph on the left and a loaded 3D structure on the right. The structure shown is Influenza A virus - Nucleoprotein (UniProt:P03466). Immunogenic regions are highlighted in dark blue on the structure. On the bottom, there is a table with the epitope information. The row is colored yellow if the displayed structure contains that epitope.

In addition, users can select their epitope of choice, which will be highlighted in green, both in the table and in the structure (Fig 3D).

Figure 3D: An epitope has been selected from the table (highlighted in green), and it is also displayed in green highlight on the 3D structure. 

Difference Between Mapped Response Frequency & Response Frequency (Normalized):

The ImmunomeBrowser is a tool that retrieves all epitopes available in the IEDB related to a given parent protein and calculates a score called the response frequency (RF). This score attempts to draw attention to more immunogenic regions of the antigen. The RF score is based on the number of positive assays at each position of the protein and uses the lower bound of the 95% confidence interval to provide a conservative estimate.

For some antigens, the range of the RF scores is small, making it challenging to see the differences in the antigen coloring. To resolve this issue, we have the option for a normalized RF, which remaps the minimum RF to 0 and the maximum to 100, to be used as a display. While this may show regions with peak RF, it may artificially imply high RF when the overall RF values are low. 

AlphaFold Predicted Models

For visualizing the 3D structure of antigens which do not have a 3D structure determined experimentally, the corresponding AlphaFold model of the antigen (if available) was retrieved from the AlphaFold Protein Structure Database. The AlphaFold models are based on the reference Uniprot sequences of proteins. After retrieving the AlphaFold model of a specific antigen, the epitope residues are mapped onto the antigen. This model, along with the epitope residues, can be visualized in the 3D viewer to give a better understanding of the spatial location of the epitope residues on the surface of the antigen.

Unusual Behavior & User Workarounds

1.  Loading Calculated Contacts in the Assays Tab 3D viewer

Please note that for larger structures with many calculated contacts (see highlighted yellow in Figure 4), the structure will take some time to load the expected interactions. You may also experience slowness when selecting and deselecting new contacts to be calculated. Do not close the pop-up or refresh the page, as it will interrupt your loading 3D viewer.

Figure 4: When the user selects to display the calculated contacts, it will take seconds to load it. It is normal to experience some lag during the loading time as all the calculations are completed. 

2. Whole white structures

When using the ImmunomeBrowser to visualize 3D antigens' structures colored by their immunogenicity, there are sometimes when the molecule is colored entirely in white, as seen in Figure 5. 

This is not an error. It can happen when all the epitopes mapped into that atomic model have a response frequency score of 0.0. To avoid this situation, the user can follow these suggestions:

• MHC assays do not contain immunogenicity information. Avoid filtering the data only for MHC assays if you want the 3D structure colored by the response frequency score in the ImmunomeBrowser.
• Filtering by specific epitopes or MHC alleles will narrow the results, which can lead to a small number of epitopes, making the structure colored white. If it is the case, try to reduce the number of filters.

Figure 5: An example of a 3D structure colored white after clicking the Map Response Frequency button. For generated this particular example, we filtered our results for MHC assays only at the IEDB homepage and selected the Hemagglutinin to be loaded in the ImmunomeBrowser.

3. How to save an image

After opening a 3D structure, the user can click on the 3-bar button symbol (1) to open the iCn3D menu, available in the top left corner of the window. To access the function to save the current canvas as a picture, the practitioner can click on File (2) -> Save File (3) -> iCn3D PNG File (4). There are some options for the resolution. We suggested saving in 4x large (5). A new window will pop up to choose the figure's name and where to save it. All the steps are shown in Figure 6.

Figure 6: Step-by-step of how any user can save an image of the loaded antigen in the iCn3D. After opening the menu (1), select file (2), save file (3), iCn3D PNG Image (4), and lastly, the user can choose any size. We recommended the 4X Large (5).